Rap1GAP-mediated MMP9 secretion is associated with poor survival in SCC

Rap1GAP functions by switching-off rap1, the ras-like protein that has been associated with carcinogenesis. Previous findings suggest that rap1GAP acts as a tumor suppressor protein in squamous cell carcinoma (SCC) by delaying the G1/S transition of the cell cycle. Furthermore, tumors derived from SCC cells transfected with rap1GAP were more invasive than corresponding controls. Matrix metalloproteinase-9 (MMP9) is an enzyme that mediates SCC invasion via degradation of the extracellular matrix. Objective: The objective of the current study was to investigate the effects of rap1GAP on invasion and progression of SCC, and the role of MMP9 in this process. Methods: Using SCC cells transfected with empty vector or rap1GAP, cell invasion and MMP secretion were determined by matrigel assays and gelatin zymography. Transcription of MMP9 was assayed by Q-RT-PCR. Immunohistochemistry was performed on oropharyngeal SCC tissue microarrays from a completed clinical study. To evaluate univariate associations between markers and ordinal variables of interest the Spearman correlation coefficient was used. Cox proportional hazards model was used to relate time-to-event outcomes to marker levels. Results: Cells transfected with rap1GAP exhibit a more invasive phenotype than corresponding vector transfected control cells. Rap1GAP upregulated transcription and secretion of MMP9. Furthermore, MMP2/MMP9 inhibitor I (Calbiochem), inhibited invasion by rap1GAP transfected cells. Immunohistochemical staining of the SCC tissue microarray, showed that rap1GAP and MMP9 expression and staining intensity are correlated (p<0.0001). The effect of MMP9 on disease-specific survival depends on N-stage (p<.05). High MMP9 is prognostic of poor survival in early N-stage and better survival in advanced N-stage lesions. Conclusion: Rap1GAP inhibits tumor growth but induces MMP9-mediated SCC invasion and tumor progression, suggesting a role for this protein as a biomarker for early N-stage, aggressive SCCs. [This work was supported by NIDCR DE16920-01, DE018512-01, UM Head and Neck SPORE developmental project grants and UM NSF-ADVANCE Crosby award].